Obesity is a major public health concern due in part to the strong associations between obesity, insulin resistance and co-morbidities of hypertension, diabetes, dyslipidemia and atherosclerosis. Our clinical investigations have focused upon patterns of fatty acid metabolism in skeletal muscle of obese individuals and the potential relation to insulin resistance and regional patterns of fat distribution. We find that in obesity, skeletal muscle has reduced efficiency for oxidation of fatty acids during fasting conditions yet essentially normal rates of uptake of plasma FFA and therefore, we postulate this leads to increased fat deposition within skeletal muscle. In cross-sectional studies, we find that a non-invasive index of fat deposition within muscle, determined using computed tomography, is at least as strong a marker of insulin resistance as visceral fat content. Unfortunately, diet induced weight loss while effective to reduce visceral and generalized adiposity, had little effect on patterns of fatty acid metabolism in skeletal muscle and did not improve the reduced activity of muscle carnitine palmitoyl transferase and other oxidative enzymes which we find in muscle of obese individuals. We posit that inefficiency in the oxidation of fatty acids, likely due to compromised transport of fatty acids into mitochondria, leads to increased fat deposition within muscle and thereby sets the conditions for lipid induced insulin resistance of obesity. In this application for renewed support, we propose to study the efficacy of exercise (4 months aerobic exercise) alone and in combination with a weight loss program (the very low calorie diet we have used previously without exercise) for improving the capacity of skeletal muscle for the utilization of fatty acids in obesity. A cohort of lean controls will be studied before and following the exercise program. At baseline and following intervention, fatty acid uptake will be measured using leg balance and stable isotope methodology together with regional indirect calorimetry studies of substrate oxidation. Insulin sensitivity will be measured using the glucose clamp method. We will continue to obtain vastos lateralis muscle by percutaneous biopsy for assay of marker enzymes of fatty acid catabolism, including plasma membrane and cytosolic fatty acid binding proteins and uncoupling protein (UCP2). In addition, we will assay ex vivo mitochondrial capacity for ATP generation from fatty acids compared to pyruvate, and assess the functional capacity of the respiratory chain to better delineate where an obesity-related impairment of fat oxidation occurs. Muscle lipid content will be ascertained by lipid extraction, by Oil Red O staining and quantitative microscopy of muscle cryosections, and by a novel, non-invasive method of water-suppressed echo planar magnetic resonance imaging of the mid-thigh. Thus, using biochemical analysis of muscle, physiologic studies of substrate kinetics, and assessments of muscle composition, we will evaluate patterns of fatty acid uptake, oxidation and storage in obesity and the efficiacy of exercise for improving fatty acid metabolism by skeletal muscle.